Method for screening an agent for preventing or ameliorating wrinkles

Abstract

There is provided a screening method which can select a photoaging-preventing or improving agent or a wrinkle-preventing or improving agent with efficiency and reliability. The screening method for a photoaging- or wrinkle-preventing or improving agent selects, as a candidate, a substance inhibiting the expression of keratin 6 and/or keratin 16 genes or keratin 6 and/or keratin 16 proteins in a skin tissue or cell.

1. A screening method for a photoaging- or wrinkle-preventing or improving agent, which selects, as a candidate, a substance inhibiting an expression of keratin 6 and/or keratin 16 genes or keratin 6 and/or keratin 16 proteins in a skin tissue or cell, wherein the skin tissue and/or cells are derived from a subject that has been exposed to UVB irradiation. 
     
     
       2. The screening method according to  claim 1 , wherein the skin tissue or cell is derived from a subject having the enhanced expression of keratin 6 and/or keratin 16 genes or keratin 6 and/or keratin 16 proteins due to an exposure to ultraviolet irradiation of 1 minimum erythema dose (MED) or lower for 4-20 consecutive weeks. 
     
     
       3. A screening method for a photoaging- or wrinkle-preventing or improving agent, comprising:
 (1) bringing a test substance into contact with a skin tissue or cell having the enhanced expression of keratin 6 and/or keratin 16 proteins; 
 (2) extracting epidermal keratin proteins from the skin tissue or cell; 
 (3) measuring expression levels of keratin 6 and/or keratin 16 proteins in the extracted epidermal keratin proteins and comparing the expression levels with expression levels of keratin 6 and/or keratin 16 proteins in a control skin tissue or cell not brought into contact with the test substance; and 
 (4) selecting the test substance decreasing the expression levels of keratin 6 and/or keratin 16 proteins as a photoaging- or wrinkle-preventing or improving agent, based on the result of the step (3), 
 wherein the skin tissue or cell are derived from a subject that has been exposed to ultraviolet radiation of 1 minimum erythema dose (MED) or lower for 4-20 consecutive weeks before (1). 
 
     
     
       4. A screening method for a photoaging- or wrinkle-preventing or improving agent, comprising:
 (1) bringing a test substance into contact with a skin tissue or cell having the enhanced expression of keratin 6 and/or keratin 16 genes; 
 (2) measuring expression levels of keratin 6 and/or keratin 16 genes in the tissue or cell brought into contact with the test substance and comparing the expression levels with expression levels of keratin 6 and/or keratin 16 genes in a control skin tissue or cell not brought into contact with the test substance; and 
 (3) selecting the test substance decreasing the expression levels of keratin 6 and/or keratin 16 genes as a photoaging- or wrinkle-preventing or improving agent, based on the result of the step (2), 
 wherein the skin tissue or cell are derived from a subject that has been exposed to ultraviolet radiation of 1 minimum erythema dose (MED) or lower for 4-20 consecutive weeks before (1). 
 
     
     
       5. The screening method of  claim 3 , further comprising measuring expression levels of keratin 1 and keratin 14 in the skin tissue or cell, wherein the expression levels of keratin 1 and keratin 14 do not change statistically significantly in the skin tissue or cell irradiated with UVB compared to the expression levels of keratin 1 and keratin 14 in a skin tissue or cell not irradiated with UVB. 
     
     
       6. The screening method of  claim 4 , further comprising measuring expression levels of keratin 1 and keratin 14 genes in the skin tissue or cell, wherein the expression levels of keratin 1 and keratin 14 genes do not change statistically significantly in the skin tissue or cell irradiated with UVB compared to the expression levels of keratin 1 and keratin 14 genes in a skin tissue or cell not irradiated with UVB. 
     
     
       7. The screening method of  claim 2 , further comprising measuring expression levels of keratin 1, keratin 10, keratin 5, and keratin 14 genes in the skin tissue or cell, wherein keratin 1, keratin 10, keratin 5, and keratin 14 do not change statistically significantly the expression levels in the skin tissue or cell irradiated with UVB compared to the expression levels of keratin 1, keratin 10, keratin 5, and keratin 14 genes in a skin tissue or cell not irradiated with UVB. 
     
     
       8. The screening method of  claim 3 , wherein the expression level of keratin 6 and/or keratin 16 protein in the UVB irradiated skin tissue or cell is increased by about 120 to 300% for keratin 6 and about 120 to 200% for keratine 16 compared to a normal skin tissue or cell. 
     
     
       9. The screening method of  claim 4 , wherein the expression level of keratin 6 and/or keratin 16 genes in the UVB irradiated skin tissue or cell is increased by about 120 to 500% compared to a normal skin tissue or cell. 
     
     
       10. The screening method of  claim 5 , wherein the expression level of keratin 6 and/or keratin 16 protein in the UVB irradiated skin tissue or cell is increased by about 120 to 300% for keratin 6 and about 120 to 200% for keratine 16 compared to a normal skin tissue or cell. 
     
     
       11. The screening method of  claim 6 , wherein the expression level of keratin 6 and/or keratin 16 genes in the UVB irradiated skin tissue or cell is increased by about 120 to 500% compared to a normal skin tissue or cell. 
     
     
       12. The screening method of  claim 1 , wherein the skin tissue or cells are derived from an epidermal portion of the skin of the subject comprising stratum corneum. 
     
     
       13. The screening method of  claim 2 , wherein the skin tissue or cells are derived from an epidermal portion of the skin of the subject comprising stratum corneum. 
     
     
       14. The screening method of  claim 3 , wherein the skin tissue or cells are derived from an epidermal portion of the skin of the subject comprising stratum corneum. 
     
     
       15. The screening method of  claim 4 , wherein the skin tissue or cells are derived from an epidermal portion of the skin of the subject comprising stratum corneum. 
     
     
       16. The screening method of  claim 5 , wherein the subject is an immunodeficient mouse comprising a human skin graft, and the skin tissue and/or cells are derived from the graft. 
     
     
       17. The screening method of  claim 6 , wherein the subject is an immunodeficient mouse comprising a human skin graft, and the skin tissue and/or cells are derived from the graft. 
     
     
       18. The screening method of  claim 7 , wherein the subject is an immunodeficient mouse comprising a human skin graft, and the skin tissue and/or cells are derived from the graft.